Lee Yu-Shang, Baratta Janie, Yu Jen, Lin Vernon W, Robertson Richard T
Department of Anatomy and Neurobiology, College of Medicine, University of California, Irvine 29697-1280, USA.
J Neurotrauma. 2002 Mar;19(3):357-67. doi: 10.1089/089771502753594927.
This study developed a slice culture model system to study axonal regeneration after spinal cord injury. This model was tested in studies of the roles of acidic fibroblast growth factor (aFGF) and peripheral nerve segments in axonal growth between pieces of spinal cord. Transverse sections of P15-P18 Sprague-Dawley rat spinal cord were collected for organotypic slice cultures. Group I consisted of two slices of spinal cord in contact with each other during the culture period. Group II consisted of two slices that were separated by 3 mm and connected by two segments of intercostal nerves. Group III consisted of single slices for studies of neuron survival. Some cultures from each group included aFGF in the culture medium. Bromodeoxyuridine (BrdU) was included in the medium for some cultures. The results showed three principal findings. First, counts of neurofilament-positive cells demonstrated that treatment with aFGF significantly increased the number of surviving neurons in culture. Second, neurofilament immunostaining and DiI tracing demonstrated axons crossing the junction between the two pieces of spinal cord or growing through the intercostal nerve segments, and these axons were seen only in cultures with aFGF treatment. Third, few cells were double stained for neurofilament and BrdU, and these were found only with aFGF treatment. These results demonstrate that (1) organotypic slice cultures present a useful model to study regeneration from spinal cord injury, (2) aFGF rescues neurons and promotes axonal growth in these cultures, and (3) segments of intercostal nerves promote axon growth between slices of spinal cord.
本研究开发了一种切片培养模型系统，用于研究脊髓损伤后的轴突再生。该模型在酸性成纤维细胞生长因子（aFGF）和周围神经节段在脊髓片段间轴突生长中的作用研究中进行了测试。收集P15 - P18 Sprague - Dawley大鼠脊髓的横切片用于器官型切片培养。第一组由培养期间相互接触的两片脊髓组成。第二组由两片相隔3毫米并由两段肋间神经连接的脊髓组成。第三组由用于神经元存活研究的单切片组成。每组的一些培养物在培养基中加入了aFGF。一些培养物的培养基中加入了溴脱氧尿苷（BrdU）。结果显示出三个主要发现。第一，神经丝阳性细胞计数表明，aFGF处理显著增加了培养物中存活神经元的数量。第二，神经丝免疫染色和DiI追踪显示轴突穿过两片脊髓之间的连接处或通过肋间神经节段生长，并且这些轴突仅在接受aFGF处理的培养物中可见。第三，很少有细胞同时被神经丝和BrdU双重染色，并且这些仅在aFGF处理时被发现。这些结果表明：（1）器官型切片培养提供了一个研究脊髓损伤后再生的有用模型；（2）aFGF拯救神经元并促进这些培养物中的轴突生长；（3）肋间神经节段促进脊髓切片间的轴突生长。
